Cyclic GMP (cGMP) is the primary intracellular messenger for the visual signaling pathway in rod and cone photoreceptor cells of the retina. Light stimulation causes the activation of the cGMP phosphodiesterase (PDE6), the central effector of this biochemical pathway, and a transient lowering of cGMP concentration that generates the electrical response. Precise regulation of the lifetime of PDE6 activation is required to control the sensitivity, amplitude and kinetics of the light response. While the basic pathway of phototransduction is well established, important aspects of the activation and regulation of PDE6 remain unknown, including a detailed understanding of how PDE6 is inhibited by its gamma subunit and how activation of PDE6 by transducin occurs. Other proteins are known to interact with PDE6 in photoreceptor cells, but their purpose is unclear. Visual impairment and retinal degenerative disease (e.g., retinitis pigmentosa) are known to result from mutations in the genes coding for PDE6 subunits, and it is also known that genetic defects in PDE6 interacting proteins also can cause retinal degeneration and/or visual dysfunction. Furthermore, the increasing therapeutic reliance on PDE5 inhibitors (e.g., Viagra) raises concerns about adverse effects on the closely related photoreceptor PDE6 enzyme and its ability to control cGMP levels during visual signaling. Advancing our understanding at the molecular level of the structure, function, regulation, and pharmacology of PDE6 will permit effective treatments to be devised to slow, halt, and (ultimately) reverse the deterioration of vision in patients afflicted with visual disorders and progressive retinal degenerative diseases resulting from genetic defects in PDE6 and its network of interacting proteins. The three specific aims of the project are to understand: (a) the sequence by which inhibition by the gamma-subunit of PDE6 is relieved when transducin activates the PDE6 holoenzyme during visual excitation;(b) how the structural differences between PDE5 and PDE6 can reveal functional differences in drug binding, allosteric regulation and functional protein expression;and (c) the network of PDE6-interacting proteins and the function they play in modulating PDE6 and the photoresponse. PUBLIC HEALTH RELEVANCE: The visual signaling pathway in rod and cone photoreceptors depends on the proper functioning and regulation of the cGMP phosphodiesterase enzyme (PDE6) during the transformation of a light stimulus into an electrical response. Genetic defects in PDE6 or therapeutic agents (e.g., erectile dysfunction drugs) that adversely affect PDE6 activity can lead to vision impairment, retinal degenerative diseases (e.g., retinitis pigmentosa, congenital stationary night-blindness), or total blindness. The proposed research will advance our understanding of the function, regulation, and pharmacology of PDE6 that will be important for developing new therapies to treat retinal diseases.